Ten Protein Kinase C (PKC) isoforms have been identified and classified into three major classes: the conventional, the novel, and the atypical PKCs. There exist differences between and within these three classes with respect to their function and expression and cell type in which they are expressed. PKC 8, n & r expression has been demonstrated in normal rat calvaria osteoblastic cells to follow a specific pattern dependent upon time in culture. In order to elucidate the possible role of PKC 5, n & 5 in osteoblastic cells differentiation, the effects of 1,25(OH)2D3 on both alkaline phosphatase and the expression of the isoforms were tested in normal rat osteoblastic cells. Cells were isolated from fetal rat calvaria by sequential collagenase digestion, cultured in BGJb medium containing 10% FCS. Cells from different days in culture were treated either with vehicle or 10 nM 1,25(OH)2D3, and char acterized for alkaline phosphatase activity using flow cytometry. PKC isoform expression was studied using anti-PKC 5, n & 5 by western blotting technique. The results demonstrate that 10 nM l(x,25(OH)2D3 had no significant effect on PKC isoform expression at days 6 and 9 of cell culture. On the other hand, at day 12 the steroid elicited a significant increase in PKC ~ and 5 expression compared to control cultures. PKCr expression at day 12 was not different from control cultures after treatment with l(x,25(OH)2D3. In addition, l(x,25(OH)2D3 significantly decreased alkaline phosphatase activity at day 12 of cell culture compared to control cells. In conclusion, our results strongly suggest that l(x,25(OH)2D3 had a differential effect on the expression of the various isoforms since no effects were observed on PKCI;. In addition l(x,25(OH)2D3 modulation of alkaline phosphatase activity inversely parallels that of PKC isoforms 8 and n expression in normal rat osteoblastic cells. Key words: Osteoblasts, Bone, Vitamin D